Vol. 28, Issue 8, 920-924, August 2000

Assessment of the Metabolic Chiral Inversion of D-Leucine in Rat by Gas Chromatography-Mass Spectrometry Combined with a Stable Isotope Dilution Analysis

Hiroshi Hasegawa, Takehisa Matsukawa, Yoshihiko Shinohara, and Takao Hashimoto

School of Pharmacy, Tokyo University of Pharmacy and Life Science, Tokyo, Japan

The stereoselective pharmacokinetics of leucine enantiomers in rats has been investigated to evaluate the inversion of D-leucine to L-enantiomer. After a bolus i.v. administration of D- or L-[²H₇]leucine to rats, blood samples were obtained over 6 h after administration and analyzed by a stereoselective gas chromatography-mass spectrometry method. Racemic [²H₃]leucine was used as an internal standard. The method involved methyl esterification and subsequent chiral derivatization with (+)--methoxy--trifluoromethylphenylacetyl chloride to form the diastereomeric amide. The derivatization made possible the separation of leucine enantiomers with good gas chromatographic behavior. Plasma concentration of both D- and L-[²H₇]leucine declined biexponentially, with elimination half-lives of 60 and 14 min, respectively. In contrast to the L-enantiomer, the D-enantiomer had a lower systemic clearance. When D-[²H₇]leucine was administered, the L-enantiomer was found to rapidly appear in plasma. About 30% of an administered dose of the D-isomer was stereospecifically inverted to the L-enantiomer. There was no measurable inversion of the L- to D-enantiomer. This methodology has made it possible to evaluate the pharmacokinetics of each enantiomer of amino acids and estimate of chiral inversion after administration of D-amino acids.